Membranous translation platforms in the chloroplast of Chlamydomonas reinhardtii.

Abstract

A small genome in chloroplasts encodes many of the polypeptide subunits of the photosynthetic electron transport complexes embedded in the membranes of thylakoid vesicles in the chloroplast stroma and synthesized by ribosomes of the bacterial-like genetic system of this semiautonomous organelle. While thylakoid membranes (TMs) are sites of translation, evidence in the unicellular alga Chlamydomonas reinhardtii supports translation on noncanonical membranes in a discrete translation zone in the chloroplast. To characterize the membranous platforms for translation and the biogenesis of TMs, we profiled membranes during chloroplast development, using the yellow-in-the-dark1 mutant, and carried out proteomic analyses on 2 membrane types proposed previously to support translation in the chloroplast of C. reinhardtii: "low-density membrane" (LDM) and "chloroplast translation membrane" (CTM). The results support the roles of LDM and CTM in the preliminary and ongoing stages of translation, respectively. Proteomics, immunoprecipitation, and transmission electron microscopy results support connections of these membranous platforms and a chloroplast envelope domain bound by cytoplasmic ribosomes. Our results contribute to a model of photosynthesis complex biogenesis in a spatiotemporal "assembly line" involving LDM and CTM as sequential stages leading to photosynthetic TMs.